Tat is an essential HIV-1 regulatory protein whose best described role is to promote high levels of viral gene expression via interactions with the HIV-1 transactivation response element (TAR) RNA (Dayton et al., 1986; Hauber et al., 1987). Full-length Tat is encoded by two exons comprising 101 amino acids (varying between 99 and 104 residues) and represents the most abundant form of Tat from patient derived HIV-1. The first exon is organized into two major domains: the activation domain, which interacts with numerous cellular proteins including cyclin T1, and the basic domain, which is primarily comprised of arginine and lysine residues. The basic domain (amino acids 49-57) is required for many of Tat's activities including nuclear localization (Hauber et al., 1989; Ruben et al., 1989) and TAR binding (Berkhout et al., 1989). The basic domain has also been reported to facilitate other Tat activities such as membrane transduction (Vivès et al., 1997), assisting HIV-1 reverse transcription (Apolloni et al., 2003) and augmenting integrin receptor binding (Barillari et al., 1993). A transdominant negative mutant is typically an altered form of a protein that can inhibit the normal function of its wild type counterpart. Engineered Tat proteins with altered basic domains possess transdominant negative phenotypes against wild type Tat. However, previous studies of Tat transdominance have used one-exon tat mutants encoding truncated proteins of 72 amino acids or less. For example, Tat truncated at residue 53 can suppress transactivation initiated by wild type Tat (Pearson et al., 1990). This is despite the mutant localizing mainly to the cytoplasm of the cell, in contrast to wild type Tat, which localizes to the nucleus. One exon tat mutants with a deleted basic domain or where the basic domain has been substituted with neutrally-charged amino acids also recapitulate the transdominant negative effects on transactivation (Orsini & Debouck, 1996; Ulich et al., 1996). Localization of Tat mutants to the nucleus, via fusion of the Tat nuclear localization signal to their carboxy termini, results in retention of the transdominant negative phenotype (Orsini & Debouck, 1996). Moreover, mutations in the activation domain of the Tat mutant, which normally suppress the transactivation function of wild type Tat, can suppress transdominance (Orsini & Debouck, 1996). The mechanism of transdominance of Tat basic-domain mutants is unclear.